1. Field of the Invention
The present invention relates to methods and test kits for the amplification and detection of human immunodeficiency virus (HIV). In particular, the present invention relates to PCR methods that use multiple primer sets to amplify all subtypes of HIV-1, including group M and group O isolates, and all subtypes of HIV-2.
2. Background Information
Progress has been made in our understanding of acquired immunodeficiency syndrome (AIDS) and its causative agent, the human immunodeficiency virus (HIV). Two groups of human immunodeficiency viruses, HIV-1 and HIV-2, have been identified. HIV-1 and HIV-2 are genetically related, but are nevertheless distinct. Both HIV-1 and HIV-2 show considerable genetic variability among their different isolates. Indeed, ten subtypes of HIV-1 (group M comprising subtypes A to I and group O comprising subtype O) have been identified. While consensus sequences have been generated for HIV-1 and HIV-2 from published nucleotide sequences of various HIV-1 and HIV-2 isolates, it is impossible to find substantial regions of absolute sequence conservation between all isolates of HIV-1 or all isolates of HIV-2. In addition to the genetic variability found among HIV isolates, many nucleotide regions of HIV are as highly conserved between HIV and non-related viruses as they are within the HIV-1 and HIV-2 families. These facts, taken together, make it extremely difficult to design polymerase chain reaction (PCR) primers and probes that will efficiently detect all group M and group O subtypes of HIV-1 and all HIV-2 subtypes without falsely detecting non-related viruses.
One of the challenges facing those attempting to develop amplification systems that detect all known HIV-1 and HIV-2 subtypes is the development of oligonucleotide primer sets which perform adequately when used together under a single set of amplification conditions (such as, salt conditions, temperatures, and amplification times). Identifying primers and detection probes that are compatible in terms of primer-primer and primer-probe interactions is an even greater challenge. Thus, in the case of the amplification and detection of HIV-1 and/or HIV-2, steps maximizing the probability of detecting highly divergent subtypes or isolates under less than ideal conditions are still needed.